Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by pathogenic autoantibodies that are dependent on T lymphocyte interactions with autoantibody producing B- lymphocytes. This interaction is reliant on CD40-1igand (CD154), which is expressed on the surface of activated CD4 T cells and which engages its cognate cell surface receptor, CD40, on B cells. Because of its critical and pleiotropic role in the immune system, CD154 expression is normally tightly regulated. Several recent reports have described dysregulated (increased and prolonged levels) expression of CD154 in patients with SLE relative to normal controls. Similar findings have been made in lupus-prone mice, and treatment of these mice with a neutralizing anti-CD154 monoclonal antibody delays and reduces the incidence of glomerulonephritis, a hallmark of SLE. However, similar approaches in humans have not proven efficacious due to unanticipated side effects involving coagulation. The expression of CD154, like that of other T cell cytokine genes, is controlled at the level of gene transcription. We have previously characterized the CD154 transcriptional promoter and demonstrated its dependence on the nuclear factor of activated T cells (NFAT) for activation-induced expression in T cells. Others have shown that CD154 expression is markedly decreased in T cells from mice deficient in NFAT1, the most prominent NFAT family member in T cells of the peripheral immune system. Therefore, we predict that NFAT1 will be critically important for endogenous CD154 expression in peripheral CD4 T cells, and that increased expression of NFATI, or other NFAT family members, contributes to the hyper-expression of CD 154 in patients with SLE. We will test the role of NFAT1 on endogenous CD 154 expression by T cells in vivo under circumstances where NFAT1 is limiting using mice that conditionally express NFATI in T cells only. To examine the role that NFAT1 and other NFAT proteins play in the abnormal expression of CD154 in SLE, primary human CD4 T cells from SLE patients and controls will be evaluated for NFAT RNA and protein levels, as well as for functional NFAT activity using reporter gene assays. Lastly, inhibition of NFAT activity in primary lupus CD4 T cells will be attempted using a newly described NFAT-specific inhibitory peptide. Ultimately, these studies may lead to alternative approaches to inhibiting NFAT activity, and the subsequent over-expression of CD154, in SLE T cells.